Isomalto-oligosaccharides syrups contain a substantial amount of branched oligo-saccharides such as isomaltose, pannose, isomaltotriose, isomaltotetraose, nigerose, kojibiose, isopanose and higher branched oligo-saccharides. The products are sold in powder or liquid form, depending on the intended application. The potential applications are situated in the food area examples are:
seasonings (mayonnaise, vinegar, soup base etc.) PA1 confectionery (candy, chewing gum, chocolate, ice cream, sherbet, syrup, pate) PA1 processed foods of fruits and vegetables (jam, marmalade, fruit sauces, pickles), meat or fish foods (ham, sausage, etc.) PA1 bakery products (bread, cake, cookie) PA1 precooked foods (salad, boiled beans, etc.) PA1 canned and bottled foods, drinks (coffee, juice, nectar, aerated drinks, lemonade, cola) PA1 convenience foods (instant coffee, instant cake base, etc.).
Isomalto-oligosaccharide syrups can further be applied as ingredients in animal feed and pet foods. Non-food application areas are cosmetics and medicine (cigarette, lipstick, toothpaste, internal medicine, etc.).
It is known for some years that isomalto-oligosaccharides are related to the increase of the general well being of humans and animals when taken orally on a regular daily basis. The main action of the oligosaccharides is to increase the numbers of bifidobacteria and lactobacilli in the large intestine and to reduce the concentration of putrifactive bacteria. Bifidobacteria are associated with some health promoting properties like the inhibition of the growth of pathogens, either by acid formation or by anti-microbial activity. They are also associated with such divers effects as the modulation of the immune system (anti-tumor properties), the reduction of the levels of triglycerides and cholesterol, the production of vitamins (B group), the reduction of blood ammonia concentrations, the prevention of translocation, the restoration of the normal gut flora after anti-microbial therapy, the production of digestive enzymes, the reduction of antibiotic associated side effects (Kohmoto T., Fukui F., Takaku H., Machida Y., et al., Bifidobacteria Microflora, 7(2)(1988),61-69; Kohmoto K., Tsuji K., Kaneko T. Shiota M., et al., Biosc. Biotech. Biochem., 56(6)(1992),937-940; Kaneko T, Kohmoto T., Kikuchi H., Fukui F., et al., Nippon Nogeikagaku Kaishi, 66(8)(1992),1211-1220, Park J-H, Jin-Young Y., Ok-Ho S., Hyun-Kyung S., et al., Kor. J. Appl. Microbiol. Biotechnol., 20(3)(1992), 237-242).
The isomalto-oligosaccharides are synthesized by a transglucosylation reaction using a D-glucosyltransferase (E.C. 2.4.1.24, transglucosidase, alpha-glucosidase). This enzyme catalyzes both hydrolytic and transfer reactions on incubation with alpha-D-gluco-oligosaccharides. The transfer occurs most frequently to 6-OH (hydroxyl group 6 of the glucose molecule), producing isomaltose from D-glucose, or panose from maltose. The enzyme can also transfer to the 2-OH or 3-OH of D-glucose to form kojibiose or nigerose, or back to 4-OH to reform maltose. As a result of transglucosidase reactions, the malto-oligosaccharides are converted into isomalto-oligosaccharides resulting in a class of oligosaccharides containing a higher proportion of glucose moieties linked by alpha-D-1,6 glucosidic linkages. The transglucosidase from A.niger acts only on oligosaccharides with a low DP (McCleary B. V., Gibson T. S., Carbohydrate Research 185(1989)147-162; Benson C. P., Kelly C.T., Fogarty W. M., J. Chem. Tech. Biotechnol., 32(1982)790-798; Pazur J. H., Tominaga Y., DeBrosse C. W., Jackman L. M., Carbohydrate Research, 61(1978) 279-290).
Isomalto-oligosaccharides can be obtained in different ways. For example glucose syrups at high dry solids concentration i.e. 60-80% are treated with glucoamylase resulting in the formation of isomalto-oligosaccharides mainly DP2. Other examples are maltose transfer achieved by addition of pullulanase to liquefied starch, branching of maltose syrups and treatment of sucrose with dextran sucrase.
Commercial production of isomalto-oligosaccharides is reported to be performed in a non-continuous manner. A normal production method (JP 61-212296, Showa Sangyo Co. Ltd.) starts with the liquefaction of a 30% ds slurry of corn, potato or tapioca starch with a thermostable alpha-amylase to a 6 to 10 DE liquefact. This liquefact is brought to pH 5 and 60.degree. C., and beta-amylase and transglucosidase are added and the saccharification is continued for 48-72 h. At the end of the saccharification period, the syrup is filtered and refined with active carbon and ion-exchangers. The pure product is finally concentrated to around 80% ds (Takaku H., Handbook of Amylases and related enzymes, Ed. The Amylase Research Society of Japan, Pergamon Press, Oxford, (1988), 215-217). The beta-amylase used originates from soy-bean or wheat, the transglucosidase mostly comes from a fungal source, preferably Aspergillus niger.
Other production methods are known, these include the conversion of starch hydrolysate with a mixture of alpha-amylase and transglucosidase (JP 41-48693, Nippon Corn Starch KK) and the conversion of starch into a high DP3 or DP4 syrup with DP3 or DP4 forming alpha-amylases in conjunction with transglucosidase to produce branched oligosaccharides (JP 31-87390, Gunei, Kagaku Kogyo).
A lot of attention has been paid to soluble enzyme systems for the production of isomalto-oligosaccharides, no such activity can be seen in the field of immobilised enzymes. Japanese patent application JP63-109790 (assigned to Showa Sangyo Co.) describes the use of an immobilised transglucosidase conjugate for the production of isomalto-oligosaccharide syrups. The conjugate is made by cross-linking of gelatin with glutaric dialdehyde in the presence of transglucosidase. The obtained conjugate has a low mechanical stability, and has to be ground before transport to the holding columns is possible. Due to the rather heterogeneous reaction the enzyme is not distributed homogeneously inside the carrier material, which leads to disturbed kinetics, yielding an end product which has not the maximally obtainable amount of isomalto-oligosaccharides. A further disadvantage is that the carrier is not reusable. After exhaustion of the enzyme activity, the whole conjugate has to be thrown away which is economically and ecologically a not preferred solution.
European patent application EP 301522 relates to the production of isomaltulose starting from a mixture of glucose and fructose. The description does not disclose the use of re-usable carriers for this mentioned process in addition the examples show only the use of non-immobilised enzymes for performing the conversion.
U.S. Pat. No. 3,935,070 relates to the isomerisation of a starch hydrolysate to convert at least a part of the dextrose to levulose. This conversion may be followed by a treatment with transglucosidase on bentonite. Bentonite is not a re-usable carrier. Moreover the starting material is a dextrose mother liquor.
Japanese patent application JP04 051899 (assigned to NGK Insulators Ltd.) discloses the use of enzymes immobilised on porous ceramic particles composed of SiO.sub.2 and MgO. These ceramic particles are not re-usable.
Japanese patent application JP62 278984 (assigned to Daikin Kogyo KK) disloses the use of a co-immobilisate composed of cells and enzymes. Also such a product is not re-usable.